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Westmead hospital

8 ARTICLES PUBLISHED IN JoVE

Biology

Multiplex PCR and Reverse Line Blot Hybridization Assay (mPCR/RLB)

Matthew V. N. O'Sullivan¹, Fei Zhou¹, Vitali Sintchenko¹, Fanrong Kong¹, Gwendolyn L. Gilbert¹

¹Centre for Infectious Diseases and Microbiology, University of Sydney

An inexpensive, high throughput method for simultaneous detection of up to 43 molecular targets is described. Applications of mPCR/RLB include microbial typing and detection of multiple pathogens from clinical samples.

Medicine

Utility of Dissociated Intrinsic Hand Muscle Atrophy in the Diagnosis of Amyotrophic Lateral Sclerosis

Parvathi Menon^1,2, Steve Vucic^1,2

¹Department of Neurology, Westmead Hospital, ²Sydney Medical School, University of Sydney, Australia

Dissociated atrophy of intrinsic hand muscles, termed the split hand, appears to be a specific feature of amyotrophic lateral sclerosis (ALS). Consequently, a novel neurodiagnostic test, termed the split hand index, was developed to quantify the clinical phenomenon of the split hand, which differentiated ALS from mimic disorders.

Immunology and Infection

Rapid Identification of Gram Negative Bacteria from Blood Culture Broth Using MALDI-TOF Mass Spectrometry

Timothy J. Gray¹, Lee Thomas¹, Tom Olma¹, David H. Mitchell¹, Jon R. Iredell^1,2,3, Sharon C. A. Chen^1,3

¹Centre for Infectious Diseases and Microbiology Laboratory Services, Institute of Clinical Pathology and Medical Research, Westmead Hospital, ²Centre for Research Excellence in Critical Infection, Westmead Millennium Institute, Westmead Hospital, ³Sydney Emerging Infectious Diseases Institute, University of Sydney, Westmead Hospital

The application of matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry (MS) directly to blood culture broth expedites the identification of bacteria. The presented method is a rapid and reliable method for identification of Gram negative bacteria directly from blood culture broth.

Medicine

Whole Genome Sequencing of Candida glabrata for Detection of Markers of Antifungal Drug Resistance

Chayanika Biswas^*1, Sharon C-A. Chen^*1,2,3, Catriona Halliday^1,2, Elena Martinez¹, Rebecca J. Rockett¹, Qinning Wang¹, Verlaine J. Timms¹, Rajat Dhakal¹, Rosemarie Sadsad¹, Karina J. Kennedy⁴, Geoffrey Playford^4,5, Deborah J. Marriott⁶, Monica A. Slavin⁷, Tania C. Sorrell^1,3, Vitali Sintchenko^1,2,3

¹Centre for Infectious Diseases and Microbiology-Public Health, Westmead Hospital, ²Centre for Infectious Diseases and Microbiology Laboratory Services, ICPMR, ³Marie Bashir Institute for Infectious Diseases and Biosecurity, The University of Sydney, ⁴Department of Microbiology and Infectious Diseases, Canberra Hospital and Health Services, Australian National University Medical School, ⁵Infection Management Services, Australian National University Medical School, ⁶Department of Microbiology and Infectious Diseases, St. Vincent's Hospital, ⁷Department of Infectious Diseases, Peter MacCallum Cancer Centre

This study implemented whole genome sequencing for analysis of mutations in genes conferring antifungal drug resistance in Candida glabrata. C. glabrata isolates resistant to echinocandins, azoles and 5-flucytosine, were sequenced to illustrate the methodology. Susceptibility profiles of the isolates correlated with presence or absence of specific mutation patterns in genes.

Environment

Ammonia Fiber Expansion (AFEX) Pretreatment of Lignocellulosic Biomass

Shishir P. S. Chundawat¹, Ramendra K. Pal¹, Chao Zhao¹, Timothy Campbell², Farzaneh Teymouri², Josh Videto², Chandra Nielson², Bradley Wieferich³, Leonardo Sousa³, Bruce E. Dale³, Venkatesh Balan⁴, Sarvada Chipkar⁵, Jacob Aguado⁵, Emily Burke⁵, Rebecca G. Ong⁵

¹Department of Chemical and Biochemical Engineering, Rutgers-State University of New Jersey, ²Michigan Biotechnology Institute (MBI), ³Department of Chemical Engineering and Materials Science, Michigan State University, ⁴Engineering Technology Department, Biotechnology Program, College of Technology, University of Houston, ⁵Department of Chemical Engineering, Michigan Technological University

Ammonia fiber expansion (AFEX) is a thermochemical pretreatment technology that can convert lignocellulosic biomass (e.g., corn stover, rice straw, and sugarcane bagasse) into a highly digestible feedstock for both biofuels and animal feed applications. Here, we describe a laboratory-scale method for conducting AFEX pretreatment on lignocellulosic biomass.

Immunology and Infection

Characterization of Human Monocyte Subsets by Whole Blood Flow Cytometry Analysis

Rekha Marimuthu^1,2, Habib Francis^1,2, Suat Dervish³, Stephen C.H. Li⁴, Heather Medbury^*1,2, Helen Williams^*1,2

¹Department of Surgery, Vascular Biology Research Centre, Westmead Hospital, ²Westmead Clinical School, Department of Surgery, The University of Sydney, ³Westmead Research Hub, Westmead Institute for Medical Research, ⁴Institute for Clinical Pathology and Medical Research, Westmead Hospital

Here we present a protocol for characterizing monocyte subsets by whole blood flow cytometry. This includes outlining how to gate the subsets and assess their expression of surface markers and giving an example of the assessment of the expression of M1 (inflammatory) and M2 markers (anti-inflammatory).

Cancer Research

An Orthotopic Resectional Mouse Model of Pancreatic Cancer

Tony C. Y. Pang^1,2,4,5, Zhihong Xu^1,2, Alpha Raj Mekapogu^1,2, Srinivasa Pothula^1,2, Therese M. Becker³, David Goldstein¹, Romano C. Pirola^1,2, Jeremy S. Wilson^1,2, Minoti V. Apte^1,2

¹Pancreatic Research Group, South Western Sydney Clinical School, University of New South Wales, ²Ingham Institute for Applied Medical Research, ³Centre for Circulating Tumour Cell Diagnostics and Research, Ingham Institute for Applied Medical Research, ⁴Surgical Innovations Unit, Westmead Hospital, ⁵Westmead Clinical School, University of Sydney

In the clinical context, patients with localized pancreatic cancer will undergo pancreatectomy followed by adjuvant treatment. This protocol reported here aims to establish a safe and effective method of modelling this clinical scenario in nude mice, through orthotopic implantation of pancreatic cancer followed by distal pancreatectomy and splenectomy.

Medicine

Delivery of Cardioactive Therapeutics in a Porcine Myocardial Infarction Model

Dinesh Selvakumar^*1,2, Emma Wilkie^*1, Tejas Deshmukh^1,2, Dhanya Ravindran¹, Yasuhito Kotake², Juntang Lu², Tony Barry², Vu Tran², Hugh Paterson³, Alfred Hing⁴, Timothy Campbell², Saurabh Kumar², Eddy Kizana^1,2, James J. H. Chong^1,2

¹Centre for Heart Research, The Westmead Institute for Medical Research, The University of Sydney, ²Department of Cardiology, Westmead Hospital, ³Sydney Imaging, Core Research Facility, The University of Sydney, ⁴Department of Cardiothoracic Surgery, Liverpool Hospital

The present protocol describes three methods of administering cardioactive therapeutic agents in a porcine model. Female landrace swine received treatment through either: (1) thoracotomy and transepicardial injection, (2) catheter-based transendocardial injection, or (3) intravenous infusion via jugular vein osmotic minipump.